mazF-mediated deletion system for large-scale genome engineering in Saccharomyces cerevisiae

A new mazF-based strategy for large-scale and scarless genome rearrangements in Saccharomyces cerevisiae was developed. We applied this method to delete designed internal (26.5 kbp) and terminal (28.9 kbp) regions located on the left arm of the chromosome XI of S. cerevisiae BY4741. The number of transformants was increased by one order of magnitude and about 90% of tested colonies were desired integrants using in vivo assembled deletion cassette containing longer flanking homology. Compared to conventional URA3 marker, in the counter-selection process, the new system generated 2–13 folds more colonies and the ratio of deletant was simultaneously elevated by 20–24%.     

Recurrent integration of papillomavirus DNA within the human 12q14–15 uterine breakpoint region in genital carcinomas

Genital carcinomas are associated with human papillomaviruses, and the viral DNA is frequently integrated in the host cell genome. Recurrent chromosomal alterations are genetic markers for specific tumor phenotypes. To demonstrate that papillomavirus DNA integration is indeed a recurrent chromosomal aberration, we mapped two independent papillomavirus integration sites in the human 12q14–15 region, one containing HPV16 DNA and the other HPV18 DNA. The two HPV integration sites map approximately 10 kbp from each other within the cosmid LLNL12NCO1–196E1 clone. The integration site corresponding to HPV16 DNA in SK-v cells is proximal to the 5′ end of a DNA segment known to be rearranged by integration of HPV18 DNA in another cervical carcinoma cell line, SW756. Both integrations are located in the PAL2 locus within the uterine leiomyoma cluster region of translocation. Genes Chromosomes Cancer 23:55–60, 1998. © 1998 Wiley-Liss, Inc.     

Differences in protein synthesized in vivo and in vitro by cells associated with the cerebral microvasculature. A protein synthesized in response to trauma?

A protein of molecular weight 71,000 (P₇₁) was synthesized by cells associated with the cerebral microvasculature in telencephalon slices incubated in vitro. This protein, however, was not synthesized in vivo. There was a 30 min delay in the synthesis of P₇₁ by brain slices. During this lag period a new ribonucleic acid species which was required for P₇₁ synthesis was produced in the slices. After this delay P₇₁ became a major product of protein synthesis. P₇₁ was also produced by microvasculature enriched fractions isolated from telencephalon.This protein may be involved in the initial response of the vascular system to injury.     

Gallid herpesvirus 1 (infectious laryngotracheitis virus): cloning and physical maps of the SA-2 strain

Summary Clones representing 90% of the genome of Gallid herpesvirus 1 (infectious laryngotracheitis virus; ILTV) were obtained and used in hybridization experiments to constructEcoRI,KpnI amdSmaI physical maps. The genome was 155 kilobase pairs (kbp) and comprised of a long unique sequence (120 kbp) and a short unique sequence (17 kbp) bounded by repeat sequences each of 9 kbp. An unrelated second pair of repeat sequences was located at 0.67 and 0.88 map untis. A terminal repeat of the unique long region (U_L) was also detected, but no isomerization of U_L was detected.     

Studies of Bacillus subtilis phage M2: a physical map of the M2 genome.

The growth characteristics and DNA synthesis of Bacillus subtilis phage M2 in sporulating cells of B. subtilis 168 were investigated and compared with phage SPO 1. Differences in both growth properties and DNA synthesis were observed at various stages of cell development. M2-DNA was analyzed by neutral and alkaline sucrose gradient centrifugation and by agarose-gel electrophoresis and was compared with 4ø-29DNA, M2-DNA is cleaved by restriction endonuclease EcoRl into five fragments, but the sizes of the DNA fragments are entirely different. M2-DNA extracted from purified phage particles contains tightly bound protein which appears to be essential for transfectivity. The four EcoRl restriction sites in M2-DNA have been mapped by analysis of partial digestion products and of fragments of the DNA-protein complex, using agarose-gel electrophoresis. The physical map of the M2 genome so obtained is compared with those of the φ29 and φ15 genomes.